AN IMPROVED MICROFLUORIMETER FOR MEASURING BRIGHTNESS OF FLUORESCENT ANTIBODY REACTIONS

Abstract

A microfluorimeter suitable for measuring fluorescence of objects as small as bacteria is described. It consists of a fluorescence microscope equipped with one photomultiplier for intermittent measurements of observed objects, and another for continuous monitoring of the excitation beam. Output of the two photomultipliers is presented on a recorder as a ratio of the two currents. The light source is a high pressure mercury arc operated on de, and the instrument is aligned and standardized with the help of a standard fluorescent crystal mounted on a microscope slide. Measurements of intestinal amebae stained with a refined fraction of fluoresceinlabeled globulin showed high reproducibility on duplicate smears in spite of the presence of gross bacterial contamination. Standard errors of the means obtained from samples of 30 amebae averaged 5.6% of the means. The technique is capable of revealing extremely fine immunological distinctions in antigenic materials like intact microorganisms, virus particles and the like, even in the presence of tissue cells and other "contaminant" debris.